To maintain homeostasis, macrophages must be capable of assuming either an inflammatory or an anti-inflammatory phenotype. To better understand the latter, we stimulated human macrophages in vitro with TLR ligands in the presence of high-density immune complexes (IC). This combination of stimuli resulted in a broad suppression of inflammatory mediators and an upregulation of molecules involved in tissue remodeling and angiogenesis. Transcriptomic analysis of TLR stimulation in the presence of IC predicted the downstream activation of AKT and the inhibition of GSK3. Consequently, we pretreated LPS-stimulated human macrophages with small molecule inhibitors of GSK3 to partially phenocopy the regulatory effects of stimulation in the presence of IC. The upregulation of DC-STAMP and matrix metalloproteases was observed on these cells and may represent potential biomarkers for this regulatory activation state. To demonstrate the presence of these anti-inflammatory, growth-promoting macrophages in a human infectious disease, biopsies from patients with leprosy (Hanseniasis) were analyzed. The lepromatous form of this disease is characterized by hypergammaglobulinemia and defective cell-mediated immunity. Lesions in lepromatous leprosy contained macrophages with a regulatory phenotype expressing higher levels of DC-STAMP and lower levels of IL-12, relative to macrophages in tuberculoid leprosy lesions. Therefore, we propose that increased signaling by FcγR cross-linking on TLR-stimulated macrophages can paradoxically promote the resolution of inflammation and initiate processes critical to tissue growth and repair. It can also contribute to infectious disease progression.

Chronic granulomatous disease (CGD) is an inborn error of immunity that is caused by defects in any 1 of the 5 subunits (gp91phox, p47phox, p22phox, p67phox, p40phox) that form the NAD phosphate oxidase complex 2 (NOX2) or in the chaperone protein essential for reactive oxygen species (ROS) that supports its assembly. These defects lead to severely reduced phagocyte-derived ROS production. Almost 50% of patients with CGD have inflammatory bowel disease (IBD) associated with dysbiosis, and the age of IBD onset may vary according to the CGD genotype. Although we previously demonstrated that the intestinal microbiota determines colitis susceptibility in CGD mice, the underlying mechanisms remain unknown. We hypothesized that NOX2 defects are associated with distinct intestinal microbiome signatures and immune responses, which impact colitis severity. Chemical colitis susceptibility was evaluated in 2 strains of CGD mice (gp91phox–/– and p47phox–/–) with distinct microbiotas from 2 different animal facilities, while also evaluating the impact of microbiota standardization and colitogenic microbiota transfer on mucosal immune responses at the intestinal barrier. Although p47phox–/– and gp91phox–/– mice that harbored a colitogenic microbiota had increased colitis severity, the intestinal epithelial cells from p47phox–/– mice produced more ROS, which was associated with increased NOX isoform gene expression. In contrast, gp91phox–/– mice had decreased mucin production and a mucosal immune response profile suggestive of increased inflammasome activation at the intestinal barrier when compared with control and p47phox–/– mice. Our findings suggest that the microbiota impacts colitis susceptibility in a CGD genotype–specific manner, thereby potentially explaining differences in the timing of IBD onset in patients with different CGD genotypes and identifying potential novel and personalized therapeutic targets.

Handling cultured isolates and clinical, environmental, or wildlife surveillance samples containing Risk Group 3 and 4 pathogens presents considerable biosafety challenges in minimizing human exposure during processing and transport. Safe handling typically requires high- or maximum-containment facilities, demanding substantial logistical planning and resources. We evaluated PrimeStore Molecular Transport Medium (PS-MTM), a guanidine-based solution created to kill pathogens and preserve nucleic acids at ambient temperatures, for inactivating Crimean-Congo hemorrhagic fever, eastern equine encephalitis, Ebola, Hendra, Japanese encephalitis, Lassa, Marburg, Nipah, Rift Valley fever, and West Nile viruses. To mimic diagnostic conditions, human whole blood spiked with any of these viruses was incubated with PS-MTM for 20-, 30-, or 60-min. Samples with titers up to 10⁷ PFU/mL exposed to PS-MTM at all time points resulted in complete loss of infectivity judged by plaque assays. A 30-min incubation provided a 50% safety margin over the minimum inactivation time and was used for quantification with the tissue culture infectious dose (TCID₅₀) assay, enabling evaluation of PS-MTM’s activity for viruses that do or do not produce well-defined plaques. Results confirmed that PS-MTM inactivated all tested viruses at titers up to 10⁷ TCID₅₀/mL, underscoring its reliability for enhancing biosafety in diagnostics, outbreak management, and surveillance.

Summary: Post COVID-19 condition (Long COVID, LC) is frequently accompanied by persistent neurological symptoms, but the mechanisms linking intestinal dysbiosis to neuroinflammation remain unclear. Here we identify gut microbiota-derived extracellular vesicles (GMEVs) as functional mediators linking LC-associated dysbiosis to systemic and neuroimmune inflammation. In a longitudinally characterized cohort, individuals with LC and neurological symptoms exhibit a persistent intestinal microbiome signature. Transplantation of LC-associated microbiota into germ-free mice induces intestinal barrier disruption and neuroinflammatory phenotypes. GMEVs from individuals with LC activate inflammasome-associated programs and impair epithelial barrier function, promote inflammatory responses in macrophages, and induce coordinated pro-inflammatory transcriptional programs in human induced pluripotent stem cell (iPSC)-derived microglia. Chronic oral administration of LC-derived GMEVs remodels the microbiota and induces intestinal and systemic inflammation with glial activation in vivo . Together, these findings support a vesicle-centered framework in which microbiota-derived extracellular vesicles translate dysbiosis into sustained immune and neuroimmune activation in a post-viral inflammatory state.